Insects have a profound impact upon agriculture and human health throughout the world. Damage and destruction due to insect activity represents, on average, a loss of 10-20% of agricultural crops, stored agricultural products, timber and livestock worldwide. In addition, quarantines imposed to control the spread of insect pests severely impinge on world trade and the import and export of agricultural products. Many of the most significant and devastating infectious diseases are transmitted to man by blood-feeding insects such as mosquitoes, flies and ticks. Fatalities associated with insect-borne disease far exceed one million annually, with associated illnesses surpassing 300 million (see, e.g., WHO Weekly Epidemiological Record, 1999, 74:265-270).
Presently, the spread and activity of agricultural product pests is chiefly controlled by the widespread application of potent, broad-spectrum chemical pesticides over agricultural fields, greenhouses, and storage facilities. Pests posing a danger to human health are targeted with the widespread spraying of insecticides in or near residential areas.
The use of conventional pesticides, however, is associated with significant hazards to the environment, human health, and non-renewable natural resources. As a result, governments throughout the world are placing increasingly severe restrictions and bans on the use of chemical pesticides. Moreover, insects develop resistance to pesticides after prolonged use, necessitating the spraying of increased levels of pesticide, or the development of new, more potent, pesticide formulations.
Thus while chemical insecticides are designed to kill insects, their non-selective effects on human health, the environment and other animal species make them damaging and controversial. As a result, there is a critical need to develop safe and effective tools to manage populations of insects that are a threat to food, resources and human health. One potential approach is to exploit knowledge of insect behavior and recent exciting advances in the molecular neurobiology of insect olfaction to develop novel strategies for insect control.
2.1. Insect Olfactory Behavior
The behavior of all animals, including humans, involves the perception of events in the environment by visual, auditory and other sensory systems and the translation of these sensory stimuli into appropriate muscle responses. In simpler organisms such as insects, the recognition of sensory stimuli results in very stereotyped or “hard-wired” behaviors. Thus, by modifying or blocking the perception of environmental cues, it is possible to alter the behavior of such animals in a predictable way. Such alterations afford a powerful means to interfere with or divert innate behaviors that have a destructive effect on human health and welfare, such as the host-finding behavior of biting insects and agricultural pests.
Many insect behaviors, such as the location and selection of mating partners, food sources and suitable places for egg laying, are driven by the recognition of specific odors in the environment. For example, the male hawkworm moth, Manduca sexta, can detect extremely low concentrations of an attractive odor, called a pheromone, produced by females of the same species, and uses this sense to pursue females over large distances (Hildebrand, 1995, Proc. Nat'l Acad. Sci. U.S.A. 92:67-74). Female navel orangeworm moths, Amyelois transitella, a pest of almonds in California, are attracted to and lay eggs on their preferred host plant in response to volatile odors emitted by almond fruits and by larvae feeding on the almonds (Curtis and Clark, 1979, Environ. Entomol. 8:330-333; Phelan et al., 1991, J. Chem. Ecol. 17:599-614). Social insects, such as ants, make extensive use of chemical cues in communication, for example in the recognition and attack of intruder ants from other colonies (Holldobler and Wilson, 1990, The Ants, Belknap Press of Harvard University Press, Cambridge, Mass.). Finally, female mosquitoes of many species, including Anopheles gambiae, the principal malaria carrier, orient toward and locate human hosts by detecting human-specific scents (Takken and Knols, 1999, Annu. Rev. Entomol. 44:131-157; Bock and Cardew, eds., 1996, Olfaction in Mosquito-Host Interactions (Ciba Foundation Symposium 200), Wiley, Chichester). Recent progress in the understanding of the molecular basis of the sense of smell provides important new insight into the mechanisms by which these odor cues elicit specific behaviors. These advances provide an exciting opportunity to develop new tools for the behavior-based control of destructive insect species.
2.2. The Molecular Biology of Insect Olfaction
Insects recognize odors in the environment using specialized olfactory organs, namely the antenna and the maxillary palps. The antenna is a highly evolved structure that extends from the head and can attain a size equivalent to the length of the organism. The maxillary palps are a pair of club-shaped structures adjacent to the proboscis. The antenna and maxillary palps are covered with tiny sensory hairs that contain nerve cells with specialized machinery that can detect odorants often at vanishingly low concentrations. The initial step in the detection of odors requires the binding of odorants to specific receptor molecules that reside on the surface of these nerve cells.
Recently, a family of roughly 60 genes encoding odorant receptors has been identified in the genome of the model insect, the fruit fly Drosophila melanogaster (Vosshall et al., 1999, Cell, 96:725-736; Clyne et al., 1999, Neuron 22:327-338; Gao and Chess, 1999, Genomics 60:31-39; Vosshall et al., 2000, Cell, 102:147-159). These odorant receptors have seven predicted transmembrane domains and belong to the large superfamily of proteins termed G-protein coupled receptors (GPCRs). The expression of 42 of these receptor genes has been detected in small, non-overlapping subsets of olfactory neurons in the antenna or maxillary palp (Vosshall et al., 2000, Cell 102:147-159). The large size of this gene family, their predicted identity as seven transmembrane domain-containing GPCRs, and their selective expression in olfactory neurons strongly implicate them in the process of olfactory recognition in the fly. More recently, functional studies (Wetzel et al., 2001, Proc. Natl. Acad. Sci. USA 98:9377-9380; Stortkuhl and Kettler, 2001, Proc. Natl. Acad. Sci. USA 98:9381-9385) have identified a candidate ligand for one of these Drosophila odorant receptor gene products, confirming their identity as receptors for behaviorally relevant odorants.
One striking exception to the rule that an individual olfactory neuron expresses a single odorant receptor gene is the odorant receptor Or83b (previously known as A45; Vosshall et al., 1999, Cell, 96:725-736), which is expressed by most, if not all, olfactory neurons in the antenna and maxillary palp. Thus, it appears that olfactory neurons actually express two odorant receptor genes: the “ubiquitous” odorant receptor gene Or83b, and one of the other “classical” odorant receptor genes.
Further molecular genetic studies in Drosophila have provided additional insight into the logic of olfactory processing in insects. How does the insect brain know what the antenna is smelling? Expression studies have revealed that individual olfactory neurons are functionally distinct in that each nerve cell expresses only one of the odorant receptor genes (Vosshall et al., 1999, Cell, 96:725-736). Olfactory neurons expressing the same receptor and therefore responsive to the same odor extend axons that converge on a fixed point in the brain (Vosshall et al., 2000, Cell, 102:147-159). Different neurons converge on different points. It immediately follows that a given odor will activate a small group of neurons in the antenna that in turn will activate distinct spatial patterns in the insect brain. The quality of a perceived odor is therefore determined by spatial patterns of activation in the brain. These patterns are then interpreted to elicit appropriate behavioral responses such as attraction, repulsion, flight, mating and feeding. Odorants that modulate such behaviors in harmful or destructive insect species will be of great value in managing populations of these harmful and destructive insects.
Citation of a reference herein shall not be construed as an admission that such reference is prior art to the present invention.